Introduction: Preeclampsia (PE) is a severe obstetrical syndrome characterized by
new-onset hypertension and proteinuria and it is often associated with fetal intrauterine
growth restriction (IUGR). PE leads to long-term health complications, so early diagnosis
would be crucial for timely prevention. There are multiple etiologies and subtypes
of PE, and this heterogeneity has hindered accurate identification in the presymptomatic
phase. Recent investigations have pointed to the potential role of small regulatory
RNAs in PE, and these species, which travel in extracellular vesicles (EVs) in the
circulation, have raised the possibility of non-invasive diagnostics. The aim of this
study was to investigate the behavior of exosomal regulatory small RNAs in the most
severe subtype of PE with IUGR.
Methods: We isolated exosomal EVs from first-trimester peripheral blood plasma samples
of women who later developed preterm PE with IUGR (n=6) and gestational age-matched
healthy controls (n=14). The small RNA content of EVs and their differential expression
were determined by next-generation sequencing and further validated by quantitative
real-time PCR. We also applied the rigorous exceRpt bioinformatics pipeline for small
RNA identification, followed by target verification and Gene Ontology analysis.
Results: Overall, >2700 small RNAs were identified in all samples and, of interest,
the majority belonged to the RNA interference (RNAi) pathways. Among the RNAi species,
16 differentially expressed microRNAs were up-regulated in PE, whereas up-regulated
and down-regulated members were equally found among the six identified Piwi-associated
RNAs. Gene ontology analysis of the predicted small RNA targets showed enrichment
of genes in pathways related to immune processes involved in decidualization, placentation
and embryonic development, indicating that dysregulation of the induced small RNAs
is connected to the impairment of immune pathways in preeclampsia development. Finally,
the subsequent validation experiments revealed that the hsa_piR_016658 piRNA is a
promising biomarker candidate for preterm PE associated with IUGR.
Discussion: Our rigorously designed study in a homogeneous group of patients unraveled
small RNAs in circulating maternal exosomes that act on physiological pathways dysregulated
in preterm PE with IUGR. Therefore, our small RNA hits are not only suitable biomarker
candidates, but the revealed biological pathways may further inform us about the complex
pathology of this severe PE subtype.
